Cytochrome P450s are important phase I metabolic
enzymes located on endoplasmic reticulum (ER) involved in the metabolism of endogenous and exogenous substances. Our previous study showed that a hepatoprotective agent
silybin restored
CYP3A expression in mouse
nonalcoholic fatty liver disease (
NAFLD). In this study we investigated how
silybin regulated P450s activity during
NAFLD. C57BL/6 mice were fed a high-fat-diet (HFD) for 8 weeks to induce
NAFLD, and were administered
silybin (50, 100 mg ·kg-1 ·d-1, i.g.) in the last 4 weeks. We showed that HFD intake induced hepatic steatosis and ER stress, leading to significant inhibition on the activity of five primary P450s including
CYP1A2,
CYP2B6,
CYP2C19,
CYP2D6, and
CYP3A in liver microsomes. These changes were dose-dependently reversed by
silybin administration. The beneficial effects of
silybin were also observed in TG-stimulated HepG2 cells in vitro. To clarify the underlying mechanism, we examined the components involved in the P450 catalytic system, membrane
phospholipids and ER membrane fluidity, and found that
cytochrome b5 (cyt b5) was significantly downregulated during ER stress, and ER membrane fluidity was also reduced evidenced by DPH polarization and lower polyunsaturated
phospholipids levels. The increased ratios of
NADP+/
NADPH and PC/PE implied Ca2+ release and disruption of cellular Ca2+ homeostasis resulted from
mitochondria dysfunction and
cytochrome c (cyt c) release. The interaction between cyt c and cyt b5 under ER stress was an important reason for P450s activity inhibition. The effect of
silybin throughout the whole course suggested that it regulated P450s activity through its anti-ER stress effect in
NAFLD. Our results suggest that ER stress may be crucial for the inhibition of P450s activity in mouse
NAFLD and
silybin regulates P450s activity by attenuating ER stress.